DESCRIPTION: This proposal is structured to test the hypothesis that regulation of the MGMT gene in mammalian cells is affected by multiple hormonal agents. This hypothesis will be tested by characterizing the cis- elements in the human MGMT promotor, the cognate proteins responsible for promotor activation, and the signalling intermediates. A variety of experimental approaches including reporter gene assay, DNA-protein interaction and in vitro transcriptions will be used to address the following specific aims: 1) To characterize steroid-dependent activation of the MGMT promotor; 2) To elucidate the mechanism of AP-1 interaction with the MGMT promotor; 3) To characterize the cAMP response element(s) and its function; 4) To identify and quantitate signalling intermediates during phorbol ester and cAMP activation, and to test whether MGMT overexpression is due to altered signalling; and 5) To clone and characterize the mouse MGMT promoter preparatory to studying MGMT regulation in vivo in wild type and transgenic mice.